A systematic study for optimal cell seeding and culture conditions in a perfusion mode bone-tissue bioreactor

Bioreactors have an important role in tissue engineering to improve cell seeding and cell culturing conditions for the production of clinically relevant bone grafts. In our study, we systematically investigated the effects of static and dynamic approaches on cell seeding and in vitro culturing of a scaffold-containing perfusion bioreactor system. For this purpose, MC3T3-E1 pre-osteoblasts were seeded on chitosan-hydroxyapatite scaffolds under different flow rates and directions. We showed that static seeding with a five-day preculture ensured higher mitochondrial activity during perfusion culture. Bidirectional perfusion culture at 0.1 and 0.5 mL/min were applied for 21 days after static seeding and 5 days of static preculture to evaluate the effects of flow rate on the viability, morphology and osteogenic differentiation of cells. We found out that 0.1 mL/min flow rate significantly increased the mitochondrial activity of the cells, whereas osteogenic differentiation could be enhanced by 0.5 mL/min flow rate. In the last part of the study, in combination with static culturing bidirectional perfusion at 0.1 mL/min was applied for cell seeding and culturing. According to the results obtained from our study, perfusion seeding followed by static culture neither increased the mitochondrial activity nor enhanced the expression of osteogenic genes of cells except for osteopontin gene. However, static seeding combined with perfusion culture enhanced cell viability and osteogenic differentiation.